Neuronal cell death is a prominent feature of many diseases, including Alzheimer's disease and amyotrophic lateral sclerosis. The long term goals of this project are to decipher the molecular mechanisms leading to apoptotic neuronal cell death. In particular, the investigators are interested in the mechanisms by which oxidative stress leads to cell death. The specific studies proposed here arise from experiments using cultures of PC12 cells and sympathetic neurons as models to probe the means by which oxidative stress causes neuronal death. An understanding of the mechanisms of these pathways would allow the design of specific therapies. The applicants will contrast and compare two different paradigms of apoptotic cell death: free radical mediated cell death and trophic factor withdrawal mediated cell death in the same neuronal cell types, PC12 cells and sympathetic neurons. By utilizing the same neuronal cells to study these two pathways they eliminate the possibility that differences in the pathways are the result of availability of different death promoting molecules. They have developed the hypotheses that nitric oxide and interleukin-1B are required for free radical mediated death to occur. They will examine these hypotheses with the following specific aims: 1. To determine how NO mediates free radical induced neuronal cell death: a. Which NOS species are necessary for cell death? b. Does NO induce p53 activity. c. Is PARP activated in free radical induced cell death? d. Is peroxynitrite a critical species in NO mediated cell death? 2. To determine how IL-1B mediates cell death induced by the down-regulation of SOD1 while it has no deleterious effects in the trophic factor deprivation paradigm. a. Does IL-1B increase NOS in PC12 cells? b. What are the IL-1 receptor levels on PC12 cells? Are they regulated in cell death? c. Are neurons from IL-1 receptor knockout mice protected from oxidative stress? d. Is IL-1B increased in bcl-2 transfected PC12 cells after V-ASOD1 treatment? These studies will enhance our understanding of how NO and IL-1B operate in apoptotic neuronal cell death. By deciphering the apoptotic death pathway we will have better insight into the treatment of diseases in which there is aberrant cell death.